Reduction of <Emphasis Type="Italic">Theileria annulata</Emphasis> Infection in Ticks Fed on Calves Immunized with Purified Larval Antigens of <Emphasis Type="Italic">Hyalomma anatolicum anatolicum</Emphasis>

Larval antigen of Hyalomma anatolicum anatolicum, the vector of Theileria annulata, was purified by two-step affinity chromatography using anti-tick gut-specific rabbit IgG and IgG from immunized cattle. The purified antigen showed the presence of a single polypeptide of 37 kDa (GHLAgP) on SDS-PAGE. Two groups (I and II) of naive crossbred calves (Bos taurus × B. indicus) were immunized with 1 mg of GHLAgP in three divided doses. Immunized calves of group I were also infected with a sublethal dose of T. annulata along with a group of non-immunized calves (group III). Animals in groups I, II, III as well a control group (group IV) were challenged with live nymphs of H. a. anatolicum on the 10th day of immunization. There was a significant reduction in the number of emerging adults of 56.9% ± 1.67% in calves of group I (p < 0.01) and 63.09% ± 1.26% in calves of group II (p < 0.001) compared to the controls. The calves of groups I and II showed antibody responses to tick antigen up to day 70 post immunization. Infection with T. annulata was determined in the salivary glands of adult ticks that developed from the nymphs used for challenge infection. In ticks taken from group I calves, there was a 75.0% ± 0.00% infection compared with only 85.0% ± 2.88% infection in ticks taken from calves of group III. Using PCR, a lower infection (83.33% ± 3.33%) was detected in ticks that developed from calves of group I compared with calves from group III (90.00% ± 2.88%). The ground-up tick supernatants (GUTS) of the ticks taken from calves of group III yielded higher infection rate and exhibited higher infectivity titre in in vitro infection assay of bovine mononuclear cells than the GUTS of the ticks taken from calves of group I. The results suggest a partial reduction in growth rate of T. annulata in ticks feeding on calves immunized with GHLAgP.     

Using indirect ELISA to assess different antigens for the serodiagnosis of Fasciola gigantica infection in cattle, sheep and donkeys

An indirect enzyme-linked immunosorbent assay (iELISA) was evaluated for its diagnostic capability in detecting antibodies against Fasciola gigantica infection in cattle, sheep and donkeys sera using crude worm, excretory–secretory and glutathione S-transferase antigens prepared from adult liver fluke. Presence of F. gigantica worms at post-mortem examination of cattle, sheep and donkey’s livers was taken as a gold standard for the evaluation of the assay. The diagnostic sensitivity, specificity and accuracy percentages of iELISA were determined for each antigen. Excretory–secretory antigen gave the best results for the serodiagnosis of F. gigantica infection in cattle, sheep and donkeys using iELISA with diagnostic sensitivity percentages of 93.3%, 94.9% and 93.3%, respectively, while the specificity percentages were 96.7%, 97.2% and 96.3%, respectively, whereas the accuracy percentages were 95%, 96% and 95.7%, respectively. The diagnostic sensitivity percentages of iELISA using crude worm antigen were 96.7%, 100% and 93.3%, respectively, while the specificity percentages were 80%, 83.3% and 85.2%, respectively, whereas the accuracy percentages were 88.3%, 86.7% and 87%, respectively. The diagnostic sensitivity percentages of iELISA using glutathione S-transferase antigen were 66.7%, 71.8% and 60%, respectively, while the specificity percentages were 70%, 77.8% and 77.8%, respectively, whereas the accuracy percentages were 68.3%, 74.7% and 73.9%, respectively. Conclusively, excretory–secretory antigen dependent iELISA can be used as a reliable serodiagnostic test for F. gigantica infection in cattle, sheep and donkeys.     

Host Switch of <Emphasis Type="Italic">Lamellodiscus elegans</Emphasis> (Monogenea: Monopisthocotylea) and <Emphasis Type="Italic">Sparicotyle chrysophrii</Emphasis> (Monogenea: Polyopisthocotylea) between Cage-reared Sparids

Sharpsnout bream (Diplodus puntazzo) has been used in Adriatic aquaculture for less than a decade, but the decreasing trend of rearing this species will probably result in its complete substitution by more exploited sea bream (Sparus aurata). Only two facilities still rear both fish species in neighbouring cages in monoculture. A switch of parasites was observed between sparids during monitoring of the gill monogeneans of farmed fish. In wild fish of the Adriatic Sea, Lamellodiscus elegans (Monogenea: Monopisthocotylea) has previously been reported in annular (Diplodus annularis), and two-banded sea bream (D. vulgaris) and sharpsnout bream (D. puntazzo), and the present study confirmed its presence also in sea bream, in low prevalence and abundance. The exclusively sea bream monogenean Sparicotyle chrysophrii (Monogenea: Polyopisthocotylea) was also isolated from sharpsnout bream, showing prevalence and abundance values even higher than in its resident host. In the occurrence of L. elegans in sea bream, the opportunistic switch resulted in lower abundance and prevalence than in the original host, while in the second case of switching the monogenean S. chrysophrii showed better reproductive capacity on a new host (sharpsnout bream). Both cases point to the possible enlargement of parasite host range.     

Co-migrating and shared antigens of selectedPasteurella haemolytica untypable strains

Bacterial cell envelope preparations from eight untypable strains ofPasteurella haemolytica were compared by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and immunoblotting with rabbit antisera prepared against the eight untypable strains (one untypable strain per rabbit) and with cattle antisera prepared againstP. haemolytica serotypes 1, 2, 5, 6, 9 and against one heterologous untypable strain. Numerous comigrating and shared antigens were recognized by the eight rabbit antisera and theP. haemolytica serotype cattle antisera. Comigrating antigens at 43 and 30 kilodaltons (kDa) were recognized by all eight rabbit antisera. Shared antigens, detected by all eight rabbit antisera when reacting againstP. haemolytica serotype 1, were recognized at 43, 32, 30, 20 and 15 kDa.     

Determination of carcinoembryonic antigen and cancer antigen (CA 15‐3) in bitches with tumours on mammary gland: preliminary report

The aim of this work was to determine levels of carcinoembryonic antigen (CEA) and cancer antigen (CA 15‐3) in the blood serum of 45 bitches. A modified procedure was used to determine the CEA and CA 15‐3 markers with the human kits using the radioimmunoassay method. Samples collected from extirpated tumour of mammary glands were histologically processed and classified as per WHO guidelines. The average age of animals with tumour was 10.00 ± 2.2 years; for healthy bitches average age was 4.2 ± 3.2 years. Values of CEA and CA 15‐3 were considered positive, if they exceeded 0.23 ng mL^?1 and 7 IU mL^?1, respectively. Average levels of CEA in the tumour group were 0.25 ± 0.06 versus 0.20 ± 0.03 in healthy bitches (P = 0.0001). The average CA 15‐3 value in bitches with tumour was 8.58 ± 1.27 versus 5.14 ± 1.34 in healthy animals (P < 0.0001).     

Genotyping of <Emphasis Type="Italic">Chlamydophila psittaci</Emphasis> strains derived from avian and mammalian species

Eleven Chlamydophila (formerly Chlamydia) psittaci strains derived from avian and mammalian species (two from dairy cows, one from duck, three from sheep, three from wild birds, and two from pigs) were identified as genotype C by outer membrane protein A gene sequencing. Genotype C had been preferentially associated with waterfowl. This paper suggests that mammals may represent an underestimated source for genotype C strains and for human psittacosis cases.     

<Emphasis Type="Italic">Theileria</Emphasis> (<Emphasis Type="Italic">Babesia</Emphasis>) <Emphasis Type="Italic">equi</Emphasis> and <Emphasis Type="Italic">Babesia caballi</Emphasis> Infections in Horses in Galicia,Spain

The control of equine piroplasmosis is becoming increasingly important to maintain the international market open to the horse industry. The purpose of this study was to demonstrate the occurrence of equine piroplasmosis (Theileria equi and Babesia caballi) in Galicia, north-west Spain, and to compare haematological and serum biochemistry parameters between non-parasitaemic horses and horses parasitaemic with T. equi and B. caballi. Sixty serum samples (control group) were taken from healthy horses pastured on two farms, and examined for evidence of equine T. equi and B. caballi infection by indirect fluorescent antibody test (IFAT). Of the 60 samples, 24 (40%) and 17 (28.3%) samples were positive for T. equi and B. caballi, respectively. Twelve (20%) samples were positive for both parasites. Haematology and serum biochemistry were compared between controls and a series of 36 horses clinically affected by T. equi (25) or B. caballi (11). Compared with the healthy group, there was a 43% and 37% decrease in the haematocrit for T. equi and B. caballi infection, respectively. Parasitaemic horses presented an intense anaemia and serum biochemistry signs of liver damage. The anaemia was more severe in T. equi-infected than in B. caballi-infected horses. Our results suggest that equine piroplasmosis is widespread in the region and is a cause for concern.     

Bordetella bronchiseptica aroA mutant as a live vaccine vehicle for heterologous porcine circovirus type 2 major capsid protein expression

Porcine circovirus type 2 (PCV2) infections cause important respiratory diseases in the pig industry and are associated with many bacterial, mycoplasmal, and viral complications. In this study, a heterologous PCV2 major capsid protein (MCP) was expressed in the Bordetella bronchiseptica aroA mutant strain (BBS-MCP) and used as a live vaccine vehicle. Mice and pigs were immunized with live BBS-MCP via the intranasal route. The antibodies against MCP were induced successfully in the serum as determined by ELISA. In the PCV2 challenge experiment, viral DNA was removed successfully from the lymph nodes of pigs vaccinated with live BBS-MCP. Overall, BBS-MCP is believed to be a good candidate for the development of a live-attenuated vaccine against PCV2 infections.     

Prevalence of <Emphasis Type="Italic">Toxoplasma gondii</Emphasis> and <Emphasis Type="Italic">Neospora caninum</Emphasis> infections in sheep from Federal District,central region of Brazil

Serum samples from 1028 sheep were collected from 32 herds within Federal District, in the central region of Brazil. The samples were examined by indirect fluorescent antibody test (IFAT) using sera diluted 1:64 and 1:50 as cut-off values for the detection of antibodies against Toxoplasma gondii and Neospora caninum, respectively. The observed prevalence for T. gondii infection was 38.22% (26.81%<CI 0.95<49.62%), and the titers ranged from 64 to 65536. The observed prevalence for N. caninum infection was 8.81% (7.08%<CI 0.95<10.53%). The titers ranged from 50 to 51200. The reactant sera to both pathogens corresponded to 4.67% of the samples. The risk factors were not determined because of the absence of negative herds for T. gondii and the high proportion of positive herds for N. caninum (87.50%). The prevalence for T. gondii infection was significantly higher among males than in females. The present work is the first report on seroprevalence of T. gondii and N. caninum in sheep from Federal District and shows that infection by both parasites is widespread in the ovine population from this region.     

Systemic responses to challenge infection withHaemonchus contortus in immune Merino sheep

Some systemic responses to single-dose infection with 10 000Haemonchus contortus infective larvae were examined in sheep already shown to have protective immunity against the parasite. The major haematological finding was a neutrophil leukocytosis that occurred after the infections became patent but not during the pre-patent period. There was no definitive eosinophilia and no discernible change in the erythrocyte parameters. Systemic hyperthermia was not conclusively evident during the pre-patent period. Enzyme-linked-immunosorbent assays (ELISA) were used to measure the secondary anti-helminth antibody response in serum during the pre-patent period when the establishment of patent infection is resisted. These ELISAs employed preparations from adult worms to represent the parasitic stages of the worm, preparations from infective larvae to represent the pre-parasitic stages of the worm, and exsheathing fluid, which is the soluble material obtained whenH. contortus larvae undergo ecdysis and transform from the pre-parasitic to the parasitic phase. Antibody responses to the three preparations differed qualitatively, indicating the presence of three different but perhaps overlapping sets of antigens. The three peaks in antibody against exsheathing fluid may reflect the pulses of antigen delivered to sheep as the parasite undergoes its three moults within the host.Abbreviations ELISA enzyme-linked immunosorbent assay - EDTA ethylene diamine tetraacetate     

Performance and nematode infection of ewe lambs on intensive rotational grazing with two different cultivars of <Emphasis Type="Italic">Panicum maximum</Emphasis>

The daily live weight gain (DLWG), faecal nematode egg counts (FEC), and packed cell volume (PCV) of Suffolk, Ile de France and Santa Inês ewe lambs were evaluated fortnightly for 56 days in the dry season (winter) and 64 days in the rainy season (summer) of 2001-2002. The animals were distributed in two similar groups, one located on Aruana and the other on Tanzania grass (Panicum maximum), in rotational grazing system at the Instituto de Zootecnia, in Nova Odessa city (SP), Brazil. In the dry season, 24 one-year-old ewe lambs were used, eight of each breed, and there was no difference (p > 0.05) between grasses for DLWG (100 g/day), although the Suffolk had higher values (p < 0.05) than the other breeds. In the rainy season, with 33 six-month-old ewe lambs, nine Suffolk, eight Ile de France and 16 Santa Inês, the DLWG was not affected by breed, but it was twice as great (71 g/day, p < 0.05) on Aruana as on Tanzania grass (30 g/day). The Santa Inês ewe lambs had the lowest FEC (p < 0.05) and the highest PCV (p < 0.05), confirming their higher resistance to Haemonchus contortus, the prevalent nematode in the rainy season. It was concluded that the best performance of ewe lambs on Aruana pastures in the rainy season is probably explained by their lower nematode infection owing to the better protein content of this grass (mean contents 11.2% crude protein in Aruana grass and 8.7% in Tanzania grass, p < 0.05) which may have improved the immunological system with the consequence that the highest PCV (p < 0.05) observed in those animals.     

The influence of age and Rhodococcus equi infection on CD1 expression by equine antigen presenting cells

There is a distinct age-associated susceptibility of horses to Rhodococcus equi infection. Initial infection is thought to occur in the neonatal and perinatal period, and only foals less than 6 months of age are typically affected. R. equi is closely related and structurally similar to Mycobacterium tuberculosis, and causes similar pathologic lesions. Protective immune responses to M. tuberculosis involve classical major histocompatibility complex (MHC)-restricted T cells that recognize peptide antigen, as well as MHC-independent T cells that recognize mycobacterial lipid antigen presented by CD1 molecules. Given the structural similarity between these two pathogens and our previous observations regarding R. equi-specific, MHC-unrestricted cytotoxic T lymphocytes (CTL), we developed 3 related hypotheses: (1) CD1 molecules are expressed on equine antigen presenting cells (APC), (2) CD1 expression on APC is less in foals compared to adults and (3) infection with live virulent R. equi induces up-regulation of CD1 on both adult and perinatal APC. CD1 expression was examined by flow cytometric analysis using a panel of monoclonal CD1 antibodies with different species and isoform specificities.

Results

Three CD1 antibodies specific for CD1b showed consistent cross reactivity with both foal and adult monocyte-derived macrophages (MDM). CD1b and MHC class II expression were significantly higher on adult MDM compared with foals. R. equi infected MDM showed significantly lower expression of CD1b, suggesting that infection with this bacterium induces down-regulation of CD1b on the cell surface. Histograms from dual antibody staining of peripheral blood mononuclear cells also revealed that 45–71% of the monocyte population stained positive for CD1b, and that the majority of these also co-expressed MHC II molecules, indicating that they were APC. The anti-CD1 antibodies showed no binding or minimal binding to bronchoalveolar lavage (BAL)-derived macrophages.

Conclusion

The CD1b isoform is evolutionarily conserved, and is present on equine MDM, as well as on circulating blood monocytes. The unique susceptibility of foals to R. equi infection may be due in part to lower expression of CD1 and MHC class II, as observed in this study. The data also suggests that infection with R. equi induces down-regulation of CD1b on equine MDM. This may represent a novel mechanism by R. equi to avoid detection and killing of infected cells by the immune system, similar to that observed when human APC are infected with M. tuberculosis.     

Sheep performance under intensive continuous grazing of native grasslands of <Emphasis Type="Italic">Paspalum notatum</Emphasis> and <Emphasis Type="Italic">Axonopus compressus</Emphasis> in the subtropical region of the Highlands of Central Mexico

Liveweight gain was evaluated in tropical Dorper X Pelibuey lambs under intensive continuous grazing of native grasslands dominated by Paspalum notatum (PN) or Axonopus compressus (AC) in the subtropics of Central Mexico. Two trials were undertaken. Trial 1 lasted 12 weeks with 10 lambs (initial weight 18 +/- 2.57 kg, 3 months old) per treatment in 2002, and Trial 2 for 13 weeks with 8 lambs (initial weight 24.0 +/- 2.0 kg, 4 months old) per treatment. Lambs were weighed once per week, and liveweight change was estimated by linear regression over day of the experiment, using individual regression coefficients as unbiased estimates of daily liveweight change; analysed in a random block design. Lambs on Trial 1 gained 0.061 kg/lamb/day on PN and 0.047 kg/lamb/day on AC (P > 0.05) at an overall mean stocking rate of 25 lambs/ha. In Trial 2, liveweight gain was significantly larger in PN (0.060 kg/lamb/day) than on AC (0.043 kg/lamb/day) (P < 0.05), at a mean stocking rate of 21.5 lambs/ha. It is concluded that intensive continuous grazing of native grasslands in the subtropics of the highlands of Central Mexico enables moderate liveweight gains for weaned lambs during the rainy season; with better results in grasslands dominated by Paspalum notatum.     

Investigation of seroprevalence of <Emphasis Type="Italic">Theileria equi</Emphasis> and <Emphasis Type="Italic">Babesia caballi</Emphasis> in horses in Nigde province,Turkey

The prevalence of equine piroplasmosis caused by Theileria equi and Babesia caballi in Nigde, in central Anatolia, Turkey has remained unknown. Serum samples were obtained from a total of 125 horses and were tested for antibodies to T. equi and B. caballi using the Indirect Fluorescence Antibody Test (IFAT). Twenty-three (18.4%) horses were seropositive for equine piroplasmosis. Anti-T. equi was observed in 16 horses (12.8%) while anti-B. caballi was detected in 12 horses (9.6%). In addition, 5 serum samples were positive for both parasites. The prevalence rates of antibodies to T. equi and B. caballi for female and male horses were statistically indifferent (p = 0.19 and 0.90). The difference between the seropositivity rates to T. equi among age groups was statistically insignificant (p = 0.44) while the difference to B. caballi among age groups is statistically significant (p = 0.01). Seropositivity rates ranged from 2.9% to 25.7% for T. equi and 2.9% to 14.3% for B. caballi from the selected districts in Nigde. A statistically significant difference on seropositivity rates for the study sites was observed for only T.equi (p = 0.03). This study indicates that T. equi is higher than B. caballi in Nigde. This study was supported by the Scientific Research Projects Unit of Nigde University (FEB 2007/08).     

Pathological,serological, and virological findings in sheep infected simultaneously with <Emphasis Type="Italic">Bluetongue</Emphasis>, <Emphasis Type="Italic">Peste-des-petits-ruminants</Emphasis>, and <Emphasis Type="Italic">Sheeppox viruses</Emphasis>

In this study, pathological, serological and virological examinations were performed on 15 sheep from a flock of 250 sheep and lambs that suffer from simultaneous naturally occurring BTV, PPRV and SPV outbreaks. SPV was diagnosed macroscopically and histopathologically, BTV was diagnosed by ELISA, and PPRV was diagnosed pathologically and by ELISA. Clinically fever, diarrhea, depression, polypnea, conjunctivitis, lacrimation, rhinitis, erosive stomatitis, edema of eyelids, photophobia, cutaneous eruption with erythematous areas especially noticeable in wool-free parts of the body and axilla lesions evolving into papules were observed. At necropsy, the most effected organs were lungs and gut. Subepicardial hemorrhages were also commonly seen. While typical pox lesions were observed in some lambs, usually fibrinous pleuropneumonia was more prominent lung lesion. SPV and PPRV lesions were seen at the histopathological examination of the lesioned tissues, BT lesions were mild than SPV and PPRV microscopically. Serum and leukocyte samples of 15 animals were examined for PPRV and BTV by ELISA; 5 samples were positive for PPRV and 6 BTV, 4 were positive for both PPRV and BTV simultaneously. One hundred animals died, most were lambs. Mortality rates were 100% in lambs and 80% in the herd.     

An Enzyme-linked Immunosorbent Assay using Immonoaffinity-purified Antigen in the Diagnosis of Caprine Paratuberculosis and Its Comparison with Conventional ELISAs

An enzyme-linked immunosorbent assay (APA-ELISA) using an immunoaffinity-purified antigen was developed and compared with the unabsorbed and absorbed ELISA procedures, using a crude antigenic preparation, for its efficacy in detecting antibodies in goat sera against Mycobacterium avium paratuberculosis. Serum samples from 89 goats belonging to three different flocks, two with a history and evidence of paratuberculosis and one without it, were subjected to each ELISA, which had been standardized on known positive sera from goats experimentally infected with paratuberculosis. Faecal culture, faecal examination and histopathology were used as indicators of infection. The diagnostic sensitivities of the unabsorbed, absorbed and APA-ELISA were 81.8%, 77.3% and 77.3% and the specificities were 90.6%, 93.7% and 96.8%, respectively. The positive predictive values of APA-ELISA (94.4%) was the highest, followed by absorbed ELISA (80.9%) and unabsorbed ELISA (72.0%). The negative predictive values for APA-ELISA, absorbed ELISA and unabsorbed ELISA were 93.0%, 92.7% and 93.8%, respectively. The results indicated the value of APA-ELISA in avoiding the need to absorb individual test sera with Mycobacterium phlei and giving more consistent results than the absorbed ELISA. The APA-ELISA was also better than the other two procedures in terms of specificity and positive predictive values.